Role of prohormone convertases in pro-neuropeptide Y processing: coexpression and in vitro kinetic investigations.

Proneuropeptide Y (ProNPY) undergoes cleavage at a single dibasic site Lys38-Arg39 resulting in the formation of 1-39 amino acid NPY which is further processed successively by carboxypeptidase-like and peptidylglycine alpha-amidating monooxygenase enzymes. To investigate whether prohormone convertases are involved in ProNPY processing, a vaccinia virus derived expression system was used to coexpress recombinant ProNPY with each of the prohormone convertases PC1/3, PC2, furin, and PACE4 in Neuro2A and NIH 3T3 cell lines as regulated neuroendocrine and constitutive prototype cell lines, respectively. The analysis of processed products shows that only PC1/3 generates NPY in NIH 3T3 cells while both PC1/3 and PC2 are able to generate NPY in Neuro2A cells. The convertases furin and PACE4 are unable to process ProNPY in either cell line. Moreover, comparative in vitro cleavage of recombinant NPY precursor by the enzymes PC1/3, PC2 and furin shows that only PC1/3 and PC2 are involved in specific cleavage of the dibasic site. Kinetic studies demonstrate that PC1/3 cleaves ProNPY more efficiently than PC2. The main difference between the cleavage efficiency is observed in the Vmax values whereas no major difference is observed in Km values. In addition the cleavage by PC1/3 and PC2 of two peptides reproducing the dibasic cleavage site with different amino acid sequence lengths namely (20-49)-ProNPY and (28-43)-ProNPY was studied. These shortened ProNPY substrates, when recognized by the enzymes, are more efficiently cleaved than ProNPY itself. The shortest peptide is not cleaved by PC2 while it is by PC1/3. On the basis of these observations it is proposed, first, that the constitutive secreted NPY does not result from the cleavage carried out by ubiquitously expressed enzymes furin and PACE4; second, that PC1/3 and PC2 are not equipotent in the cleavage of ProNPY; and third, substrate peptide length might discriminate PC1/3 and PC2 processing activity.

3-year follow-up results of bone mineral content and density after a school-based physical activity randomized intervention trial.

BACKGROUND: As an important modifiable lifestyle factor in osteoporosis prevention, physical activity has been shown to positively influence bone mass accrual during growth. We have previously shown that a nine month general school based physical activity intervention increased bone mineral content (BMC) and density (aBMD) in primary school children. From a public health perspective, a major key issue is whether these effects persist during adolescence. We therefore measured BMC and aBMD three years after cessation of the intervention to investigate whether the beneficial short-term effects persisted. METHODS: All children from 28 randomly selected first and fifth grade classes (intervention group (INT): 16 classes, n=297; control group (CON): 12 classes, n=205) who had participated in KISS (Kinder-und Jugendsportstudie) were contacted three years after cessation of the intervention program. The intervention included daily physical education with daily impact loading activities over nine months. Measurements included anthropometry, vigorous physical activity (VPA) by accelerometers, and BMC/aBMD for total body, femoral neck, total hip, and lumbar spine by dual-energy X-ray absorptiometry (DXA). Sex- and age-adjusted Z-scores of BMC or aBMD at follow-up were regressed on intervention (1 vs. 0), the respective Z-score at baseline, gender, follow-up height and weight, pubertal stage at follow-up, previous and current VPA, adjusting for clustering within schools. RESULTS: 377 of 502 (75%) children participated in baseline DXA measurements and of those, 214 (57%) participated to follow-up. At follow-up INT showed significantly higher Z-scores of BMC at total body (adjusted group difference: 0.157 units (0.031-0.283); p=0.015), femoral neck (0.205 (0.007-0.402); p=0.042) and at total hip (0.195 (0.036 to 0.353); p=0.016) and higher Z-scores of aBMD for total body (0.167 (0.016 to 0.317); p=0.030) compared to CON, representing 6-8% higher values for children in the INT. No differences could be found for the remaining bone parameters. For the subpopulation with baseline VPA (n=163), effect sizes became stronger after baseline VPA adjustment. After adjustment for baseline and current VPA (n=101), intervention effects were no longer significant, while effect sizes remained the same as without adjustment for VPA. CONCLUSION: Beneficial effects on BMC of a nine month general physical activity intervention appeared to persist over three years. Part of the maintained effects may be explained by current physical activity.

Age-related changes in the bimanual advantage and in brain oscillatory activity during tapping movements suggest a decline in processing sensory reafference

Deficits in the processing of sensory reafferences have been suggested as accounting for age-related decline in motor coordination. Whether sensory reafferences are accurately processed can be assessed based on the bimanual advantage in tapping: because of tapping with an additional hand increases kinesthetic reafferences, bimanual tapping is characterized by a reduced inter-tap interval variability than unimanual tapping. A suppression of the bimanual advantage would thus indicate a deficit in sensory reafference. We tested whether elderly indeed show a reduced bimanual advantage by measuring unimanual (UM) and bimanual (BM) self-paced tapping performance in groups of young (n = 29) and old (n = 27) healthy adults. Electroencephalogram was recorded to assess the underlying patterns of oscillatory activity, a neurophysiological mechanism advanced to support the integration of sensory reafferences. Behaviorally, there was a significant interaction between the factors tapping condition and age group at the level of the inter-tap interval variability, driven by a lower variability in BM than UM tapping in the young, but not in the elderly group. This result indicates that in self-paced tapping, the bimanual advantage is absent in elderly. Electrophysiological results revealed an interaction between tapping condition and age group on low beta band (14âeuro"20 Hz) activity. Beta activity varied depending on the tapping condition in the elderly but not in the young group. Source estimations localized this effect within left superior parietal and left occipital areas. We interpret our results in terms of engagement of different mechanisms in the elderly depending on the tapping mode: a âeuro~kinestheticâeuro? mechanism for UM and a âeuro~visual imageryâeuro? mechanism for BM tapping movement.

Recent advances in remote sensing image processing

Remote sensing image processing is nowadays a mature research area. The techniques developed in the field allow many real-life applications with great societal value. For instance, urban monitoring, fire detection or flood prediction can have a great impact on economical and environmental issues. To attain such objectives, the remote sensing community has turned into a multidisciplinary field of science that embraces physics, signal theory, computer science, electronics, and communications. From a machine learning and signal/image processing point of view, all the applications are tackled under specific formalisms, such as classification and clustering, regression and function approximation, image coding, restoration and enhancement, source unmixing, data fusion or feature selection and extraction. This paper serves as a survey of methods and applications, and reviews the last methodological advances in remote sensing image processing.

Ultrastructural, morphometrical and immunocytochemical analyses of the exocrine pancreas in a hibernating dormouse.

Pancreatic acinar cells of euthermic, hibernating and arousing individuals of the hazel dormouse Muscardinus avellanarius (Gliridae) have been observed at the electron-microscopic level and analysed by means of ultrastructural morphometry and immunocytochemistry in order to investigate possible fine structural changes of cellular components during periods of strikingly different degrees of metabolic activity. During hibernation, the cisternae of the rough endoplasmic reticulum (RER) flatten assuming a parallel pattern, the Golgi apparatus is extremely reduced and the mitochondria contain many electron-dense particles. The cell nuclei appear irregularly shaped, with deep indentations containing small zymogen granules. They also contain abundant coiled bodies and unusual constituents, such as amorphous bodies and dense granular bodies. Large numbers of zymogen granules occur in all animals. However, the acinar lumina are open and filled with zymogen only in euthermic animals, whereas, in hibernating and arousing individuals, they appear to be closed. Morphometrical analyses indicate that, in pancreatic acinar cells, nuclei and zymogen granules significantly decrease in size from euthermia to hibernation, probably reflecting a drastic decrease of metabolic activities, mainly protein synthesis and processing. In all the studied animals, immunocytochemistry with specific antibodies has revealed an increasing gradient in alpha-amylase content along the RER-Golgi-zymogen granule pathway, reflecting the protein concentration along the secretory pathway. Moreover, during deep hibernation, significantly larger amounts of alpha-amylase accumulate in RER and zymogen granules in comparison to the other seasonal phases analysed. Upon arousal, all cytoplasmic and nuclear constituents restore their euthermic aspect and all morphometrical and immunocytochemical parameters exhibit the euthermic values, thereby indicating a rapid resumption of metabolic activities.

Deoxyribonucleic acid content as an indicator of progression of squamous cell carcinogenesis in the esophagus: comparative analysis on imprint-cytospin and tissue section preparation.

BACKGROUND: The purpose of this study was to explore the potential use of image analysis on tissue sections preparation as a predictive marker of early malignant changes during squamous cell (SC) carcinogenesis in the esophagus. Results of DNA ploidy quantification on formalin-fixed, paraffin-embedded tissue using two different techniques were compared: imprint-cytospin and 6 microm thick tissue sections preparation. METHODS: This retrospective study included 26 surgical specimens of squamous cell carcinoma (SCC) from patients who underwent surgery alone at the Department of Surgery in CHUV Hospital in Lausanne between January 1993 and December 2000. We analyzed 53 samples of healthy tissue, 43 tumors and 7 lymph node metastases. RESULTS: Diploid DNA histogram patterns were observed in all histologically healthy tissues, either distant or proximal to the lesion. Aneuploidy was observed in 34 (79%) of 43 carcinomas, namely 24 (75%) of 32 early squamous cell carcinomas and 10 (91%) of 11 advanced carcinomas. DNA content was similar in the different tumor stages, whether patients presented with single or multiple synchronous tumors. All lymph node metastases had similar DNA content as their primary tumor. CONCLUSIONS: Early malignant changes in the esophagus are associated with alteration in DNA content, and aneuploidy tends to correlate with progression of invasive SCC. A very good correlation between imprint-cytospin and tissue section analysis was observed. Although each method used here showed advantages and disadvantages; tissue sections preparation provided useful information on aberrant cell-cycle regulation and helped select the optimal treatment for the individual patient along with consideration of other clinical parameters.

Techniques for CT and MR, post-processing, radiation.

Brain perfusion can be assessed by CT and MR. For CT, two major techniquesare used. First, Xenon CT is an equilibrium technique based on a freely diffusibletracer. First pass of iodinated contrast injected intravenously is a second method,more widely available. Both methods are proven to be robust and quantitative,thanks to the linear relationship between contrast concentration and x-ray attenuation.For the CT methods, concern regarding x-ray doses delivered to the patientsneed to be addressed. MR is also able to assess brain perfusion using the firstpass of gadolinium based contrast agent injected intravenously. This method hasto be considered as a semi-quantitative because of the non linear relationshipbetween contrast concentration and MR signal changes. Arterial spin labelingis another MR method assessing brain perfusion without injection of contrast. Insuch case, the blood flow in the carotids is magnetically labelled by an externalradiofrequency pulse and observed during its first pass through the brain. Eachof this various CT and MR techniques have advantages and limits that will be illustratedand summarised.Learning Objectives:1. To understand and compare the different techniques for brain perfusionimaging.2. To learn about the methods of acquisition and post-processing of brainperfusion by first pass of contrast agent for CT and MR.3. To learn about non contrast MR methods (arterial spin labelling).

Characterization of OsMIK in a rice mutant with reduced phytate content reveals an insertion of a rearranged retrotransposon.

The rice low phytic acid (lpa) mutant Os-lpa-XS110-1(XS-lpa) has ~45 % reduction in seed phytic acid (PA) compared with the wild-type cultivar Xiushui 110. Previously, a single recessive gene mutation was shown to be responsible for the lpa phenotype and was mapped to a region of chromosome 3 near OsMIK (LOC_Os03g52760) and OsIPK1 (LOC_Os03g51610), two genes involved in PA biosynthesis. Here, we report the identification of a large insert in the intron of OsMIK in the XS-lpa mutant. Sequencing of fragments amplified through TAIL-PCRs revealed that the insert was a derivative of the LINE retrotransposon gene LOC_Os03g56910. Further analyses revealed the following characteristics of the insert and its impacts: (1) the inserted sequence of LOC_Os03g56910 was split at its third exon and rejoined inversely, with its 5' and 3' flanking sequences inward and the split third exon segments outward; (2) the LOC_Os03g56910 remained in its original locus in XS-lpa, and the insertion probably resulted from homologous recombination repair of a DNA double strand break; (3) while the OsMIK transcripts of XS-lpa and Xiushui 110 were identical, substantial reductions of the transcript abundance (~87 %) and the protein level (~60 %) were observed in XS-lpa, probably due to increased methylation in its promoter region. The above findings are discussed in the context of plant mutagenesis, epigenetics and lpa breeding.

Myelin basic protein content of aggregating rat brain cell cultures treated with cytokines and/or demyelinating antibody: effects of macrophage enrichment.

The demyelinative potential of the cytokines interleukin-1 alpha (IL-1 alpha), interferon-gamma (IFN-gamma), and tumor necrosis factor-alpha (TNF-alpha) has been investigated in myelinating aggregate brain cell cultures. Treatment of myelinated cultures with these cytokines resulted in a reduction in myelin basic protein (MBP) content. This effect was additively increased by anti-myelin/oligodendrocyte glycoprotein (alpha-MOG) in the presence of complement. Qualitative immunocytochemistry demonstrated that peritoneal macrophages, added to the fetal telencephalon cell suspensions at the start of the culture period, successfully integrated into aggregate cultures. Supplementing the macrophage component of the cultures in this fashion resulted in increased accumulation of MBP. The effect of IFN-gamma on MBP content of cultures was not affected by the presence of macrophages in increased numbers.